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PloS One 2014Albuminuria is a strong, independent predictor of chronic kidney disease progression. We hypothesize that podocyte processing of albumin via the lysosome may be an...
Albuminuria is a strong, independent predictor of chronic kidney disease progression. We hypothesize that podocyte processing of albumin via the lysosome may be an important determinant of podocyte injury and loss. A human urine derived podocyte-like epithelial cell (HUPEC) line was used for in vitro experiments. Albumin uptake was quantified by Western blot after loading HUPECs with fluorescein-labeled (FITC) albumin. Co-localization of albumin with lysosomes was determined by confocal microscopy. Albumin degradation was measured by quantifying FITC-albumin abundance in HUPEC lysates by Western blot. Degradation experiments were repeated using HUPECs treated with chloroquine, a lysosome inhibitor, or MG-132, a proteasome inhibitor. Lysosome activity was measured by fluorescence recovery after photo bleaching (FRAP). Cytokine production was measured by ELISA. Cell death was determined by trypan blue staining. In vivo, staining with lysosome-associated membrane protein-1 (LAMP-1) was performed on tissue from a Denys-Drash trangenic mouse model of nephrotic syndrome. HUPECs endocytosed albumin, which co-localized with lysosomes. Choloroquine, but not MG-132, inhibited albumin degradation, indicating that degradation occurs in lysosomes. Cathepsin B activity, measured by FRAP, significantly decreased in HUPECs exposed to albumin (12.5% of activity in controls) and chloroquine (12.8%), and declined further with exposure to albumin plus chloroquine (8.2%, p<0.05). Cytokine production and cell death were significantly increased in HUPECs exposed to albumin and chloroquine alone, and these effects were potentiated by exposure to albumin plus chloroquine. Compared to wild-type mice, glomerular staining of LAMP-1 was significantly increased in Denys-Drash mice and appeared to be most prominent in podocytes. These data suggest lysosomes are involved in the processing of endocytosed albumin in podocytes, and lysosomal dysfunction may contribute to podocyte injury and glomerulosclerosis in albuminuric diseases. Modifiers of lysosomal activity may have therapeutic potential in slowing the progression of glomerulosclerosis by enhancing the ability of podocytes to process and degrade albumin.
Topics: Albumins; Animals; Cells, Cultured; Denys-Drash Syndrome; Disease Models, Animal; Endocytosis; Fluorescein-5-isothiocyanate; Humans; Lysosomes; Mice; Mice, Transgenic; Podocytes; Proteolysis; Repressor Proteins; Serum Albumin; WT1 Proteins
PubMed: 24924335
DOI: 10.1371/journal.pone.0099771 -
Human Genetics Apr 1993The direct involvement of the Wilm's tumor suppressor gene (WT1) in Denys-Drash syndrome through mutations within exons 8 or 9 has recently been established. The absence...
The direct involvement of the Wilm's tumor suppressor gene (WT1) in Denys-Drash syndrome through mutations within exons 8 or 9 has recently been established. The absence of such alterations in three patients with Frasier syndrome provides a molecular basis for distinguishing these two syndromes that are associated with streak gonads, pseudohermaphroditism and renal failure.
Topics: Base Sequence; DNA, Single-Stranded; Female; Genes, Wilms Tumor; Gonadal Dysgenesis; Humans; Kidney Failure, Chronic; Molecular Sequence Data; Polymerase Chain Reaction; Syndrome; Wilms Tumor
PubMed: 8386697
DOI: 10.1007/BF00218274 -
CEN Case Reports May 2023Wilms tumor 1 (WT1) is the causative gene of Denys-Drash syndrome and Frasier syndrome, and in most cases, kidney failure develops after birth. We report an unusual case...
Wilms tumor 1 (WT1) is the causative gene of Denys-Drash syndrome and Frasier syndrome, and in most cases, kidney failure develops after birth. We report an unusual case of Potter sequence due to fetal nephropathy and kidney failure with a WT1 mutation. The neonate was born at 37 weeks of gestation, and had no distinctive facial appearance or anomalies of the extremities. The external genitalia were ambiguous. Presence of a penile-like structure or hypertrophic clitoris was noted, and the urethra opened at the base of the penis or clitoris. On ultrasonographic examination, the kidney sizes were small. No kidney cysts were noted, but the kidney parenchymal luminosity was increased. Although the neonate received mechanical ventilation because of severe retractive breathing after birth, he died of poor oxygenation due to air leak syndrome at 60 h after birth. The congenital anomalies of the kidney and urinary tract (CAKUT) gene panel revealed a heterozygous missense mutation in WT1 [NM_024426.6:exon9:c.1400G > A, p.(Arg467Gln)]. In WT1, missense mutations are associated with earlier onset of nephropathy than nonsense or splicing mutations. However, severe cases of fetal onset and early neonatal death with WT1 mutations are rare, and only one severe case with the same missense mutation in WT1 has been reported. Therefore, WT1 mutation may be suspected in Potter sequence patients with external genital abnormalities, and the WT1 missense mutation in our case [NM_024426.6:exon9:c.1400G > A, p.(Arg467Gln)] may indicate a severe case with fetal onset of nephropathy and kidney failure.
Topics: Male; Infant, Newborn; Humans; WT1 Proteins; Wilms Tumor; Mutation; Kidney Neoplasms; Renal Insufficiency
PubMed: 36227513
DOI: 10.1007/s13730-022-00742-x -
Proceedings of the National Academy of... Mar 1999The Wilms tumor-suppressor gene, WT1, plays a key role in urogenital development, and WT1 dysfunction is implicated in both neoplastic (Wilms tumor, mesothelioma,...
The Wilms tumor-suppressor gene, WT1, plays a key role in urogenital development, and WT1 dysfunction is implicated in both neoplastic (Wilms tumor, mesothelioma, leukemias, and breast cancer) and nonneoplastic (glomerulosclerosis) disease. The analysis of diseases linked specifically with WT1 mutations, such as Denys-Drash syndrome (DDS), can provide valuable insight concerning the role of WT1 in development and disease. DDS is a rare childhood disease characterized by a nephropathy involving mesangial sclerosis, XY pseudohermaphroditism, and/or Wilms tumor (WT). DDS patients are constitutionally heterozygous for exonic point mutations in WT1, which include mutations predicted to truncate the protein within the C-terminal zinc finger (ZF) region. We report that heterozygosity for a targeted murine Wt1 allele, Wt1(tmT396), which truncates ZF3 at codon 396, induces mesangial sclerosis characteristic of DDS in adult heterozygous and chimeric mice. Male genital defects also were evident and there was a single case of Wilms tumor in which the transcript of the nontargeted allele showed an exon 9 skipping event, implying a causal link between Wt1 dysfunction and Wilms tumorigenesis in mice. However, the mutant WT1(tmT396) protein accounted for only 5% of WT1 in both heterozygous embryonic stem cells and the WT. This has implications regarding the mechanism by which the mutant allele exerts its effect.
Topics: Animals; Base Sequence; DNA-Binding Proteins; Genes, Wilms Tumor; Genetic Linkage; Genitalia, Male; Humans; Kidney Diseases; Male; Mice; Molecular Sequence Data; Mutation; Syndrome; Transcription Factors; WT1 Proteins; Zinc Fingers
PubMed: 10077614
DOI: 10.1073/pnas.96.6.2931 -
Endocrine Journal Dec 2000Denys-Drash syndrome (DDS) is characterized by genital anomaly, early onset nephropathy and high risk for developing Wilms' tumor (WT). Recently, mutations in exon 8 or...
Denys-Drash syndrome (DDS) is characterized by genital anomaly, early onset nephropathy and high risk for developing Wilms' tumor (WT). Recently, mutations in exon 8 or 9 of the Wilms' tumor suppressor gene (WT1) have been found in the majority of DDS patients studied. We analyzed these two exons of the WT1 gene in genomic DNA from two female patients with DDS by using polymerase-chain reaction (PCR) and direct sequencing. The patients were accompanied with normal external genitalia, early onset renal failure between 6 and 12 months of age, and unilateral Wilms' tumor. Genomic DNA was isolated from peripheral blood leucocytes of the patients. Amplification of exons 8 and 9 of the WT1 gene by PCR was performed, and direct sequencing of the PCR product was performed using an automatic DNA sequencer. Two heterozygous missense mutations were found in these patients, including a missense mutation in exon 9 at codon 388 replacing the wild-type Cys with Phe, and a previously described mutation in exon 9 at codon 398 replacing the wild-type Leu with Pro. Cys388Phe is a novel mutation in the WT1 gene in the DDS. These cases are considered to be "incomplete DDS" with nephropathy and Wilms' tumor and without genital anomaly, the validity of which has been confirmed by mutation analysis.
Topics: Adolescent; Base Sequence; Child, Preschool; Codon; DNA Mutational Analysis; Exons; Female; Genes, Wilms Tumor; Genitalia, Female; Heterozygote; Humans; Kidney Diseases; Mutation, Missense; Polymerase Chain Reaction; Risk Factors; Sequence Analysis, DNA; Syndrome; Wilms Tumor
PubMed: 11228042
DOI: 10.1507/endocrj.47.683 -
Journal of Indian Association of... Oct 2014To evaluate the outcome of children with bilateral Wilms' tumor (BWT) treated on All India Institute of Medical Sciences-Wilms Tumor-99 (AIIMS-WT-99) protocol.
PURPOSE
To evaluate the outcome of children with bilateral Wilms' tumor (BWT) treated on All India Institute of Medical Sciences-Wilms Tumor-99 (AIIMS-WT-99) protocol.
MATERIALS AND METHODS
All children with BWT, registered in our solid tumor clinic from August 1999 through December 2010 were included.
RESULTS
Of the 178 fresh cases of Wilms Tumor (WT) treated during this period, 11 (6.2%) had bilateral involvement. All patients except one (12 and 3 cm), had massive bilateral tumors of more than 10 cm on each side. There were eight boys and three girls in the age range 6-30 months. One patient had Denys-Drash syndrome. Twenty renal units were operated upon (12 tumorectomy, five partial nephrectomy, and three nephrectomies), while one patient with inferior vena cava (IVC) thrombus died of renal failure. Tumor spill occurred in three units, lymphnode was positive in two patients. Local recurrence occurred in four patients (six of 18 renal units (33%)-two bilateral and two unilateral). There was one recurrence in the liver that was treated with radio-frequency ablation. The 5-year overall survival (OS) was 90% (95% confidence interval (CI) = 50.8-98.6) and the relapse free survival (RFS) was 38% (95% CI = 6.1-71.6).
CONCLUSION
Massive BWT respond poorly to preoperative chemotherapy, are often not amenable to partial nephrectomy/tumorectomy and have a higher local recurrence rate, giving a poor RFS.
PubMed: 25336802
DOI: 10.4103/0971-9261.142005 -
CMAJ : Canadian Medical Association... Aug 2016Rare diseases often present in the first days and weeks of life and may require complex management in the setting of a neonatal intensive care unit (NICU). Exhaustive...
BACKGROUND
Rare diseases often present in the first days and weeks of life and may require complex management in the setting of a neonatal intensive care unit (NICU). Exhaustive consultations and traditional genetic or metabolic investigations are costly and often fail to arrive at a final diagnosis when no recognizable syndrome is suspected. For this pilot project, we assessed the feasibility of next-generation sequencing as a tool to improve the diagnosis of rare diseases in newborns in the NICU.
METHODS
We retrospectively identified and prospectively recruited newborns and infants admitted to the NICU of the Children's Hospital of Eastern Ontario and the Ottawa Hospital, General Campus, who had been referred to the medical genetics or metabolics inpatient consult service and had features suggesting an underlying genetic or metabolic condition. DNA from the newborns and parents was enriched for a panel of clinically relevant genes and sequenced on a MiSeq sequencing platform (Illumina Inc.). The data were interpreted with a standard informatics pipeline and reported to care providers, who assessed the importance of genotype-phenotype correlations.
RESULTS
Of 20 newborns studied, 8 received a diagnosis on the basis of next-generation sequencing (diagnostic rate 40%). The diagnoses were renal tubular dysgenesis, SCN1A-related encephalopathy syndrome, myotubular myopathy, FTO deficiency syndrome, cranioectodermal dysplasia, congenital myasthenic syndrome, autosomal dominant intellectual disability syndrome type 7 and Denys-Drash syndrome.
INTERPRETATION
This pilot study highlighted the potential of next-generation sequencing to deliver molecular diagnoses rapidly with a high success rate. With broader use, this approach has the potential to alter health care delivery in the NICU.
Topics: Female; Genetic Association Studies; Genetic Testing; High-Throughput Nucleotide Sequencing; Humans; Infant, Newborn; Intensive Care Units, Neonatal; Male; Mutation; Ontario; Pilot Projects; Prospective Studies; Rare Diseases; Retrospective Studies
PubMed: 27241786
DOI: 10.1503/cmaj.150823 -
Kidney International Dec 2016The WT1 (Wilm's tumor suppressor) gene is expressed throughout life in podocytes and is essential for the functional integrity of the glomerular filtration barrier....
The WT1 (Wilm's tumor suppressor) gene is expressed throughout life in podocytes and is essential for the functional integrity of the glomerular filtration barrier. We have previously shown that CMIP (C-Maf inducing protein) is overproduced in podocyte diseases and alters intracellular signaling. Here we isolated the proximal region of the human CMIP promoter and showed by chromatin immunoprecipitation assays and electrophoretic-mobility shift that Wilm's tumor protein (WT1) bound to 2 WT1 response elements, located at positions -290/-274 and -57/-41 relative to transcription start site. Unlike the human CMIP gene, only one Wt1 response element was identified in the mouse Cmip proximal promoter located at position -217/-206. Luciferase reporter assays indicated that WT1 dose-dependently inhibited the transcriptional induction of the CMIP promoter. Transfection of decoy oligonucleotides mimicking the WT1 response elements prevented the inhibition of WT1 on CMIP promoter activity. Furthermore, WT1 silencing promoted Cmip expression. In line with these findings, the abundance of Cmip was early and significantly increased at the transcript and protein level in podocytes displaying a primary defect in Wt1, including Denys-Drash syndrome and Frasier syndrome. Thus, WT1 is a major repressor of the CMIP gene in physiological situations, while conditional deletion of CMIP in the developing kidney did not affect the development of mature glomeruli.
Topics: Adaptor Proteins, Signal Transducing; Animals; Base Sequence; Denys-Drash Syndrome; Female; Frasier Syndrome; Gene Expression Regulation; Humans; Kidney; Male; Mice; Podocytes; Promoter Regions, Genetic; WT1 Proteins
PubMed: 27650733
DOI: 10.1016/j.kint.2016.07.016 -
Journal of Indian Association of... 2019Synchronous Bilateral Wilms tumor (sBWT).
CONTEXT
Synchronous Bilateral Wilms tumor (sBWT).
AIMS
This study aimed to assess the outcome of patients with sBWT treated on SIOP protocol.
SETTINGS AND DESIGN
Retrospective and prospective randomized study.
SUBJECTS AND METHODS
SIOP 93-01 protocol was used to study nine patients of sBWT in a single center and followed up over a period from 2 to 5 years.
STATISTICAL ANALYSIS USED
Unpaired -test and Mann-Whitney U-test were used for analysis.
RESULTS
Of nine patients, six were included in the study as three patients lost to follow-up. Among the six patients, there were four girls and two boys with a median age of 2 years. Mean regression in the size of tumor was 87% in four out of six patients. Tumor with unfavorable histology showed 32% response (ratio of favorable: unfavorable histology 2:1). Event-free survival rate was 81.3% and overall survival was 90% over 2-5 years. Recurrence was seen in two patients of whom one had Denys-Drash syndrome. Mean DTPA glomerular filtration rate was 91.4/ml/min/1.73 m preoperatively and that of 3 months after completion of treatment was 84/ml/min/1.73 m. Health-related quality of life (HRQOL) using Pediatric Quality of Life Inventory and Lansky Play Performance Scale revealed significant improvement results of all functioning domains such as physical, social, emotional, and school subscales with < 0.05 and performance scale ( < 0.04).
CONCLUSIONS
We suggest SIOP protocol for sBWT and bilateral nephron-sparing surgery in two stages. However, long-term follow-up is required to assess the ultimate renal function outcome. HRQOL is an essential guide in improving the conditions of pediatric cancer survivors.
PubMed: 30686888
DOI: 10.4103/jiaps.JIAPS_42_18 -
Cancers Oct 2021(1) Background: about 10% of Wilms Tumor (WT) patients have a malformation or cancer predisposition syndrome (CPS) with causative germline genetic or epigenetic...
(1) Background: about 10% of Wilms Tumor (WT) patients have a malformation or cancer predisposition syndrome (CPS) with causative germline genetic or epigenetic variants. Knowledge on CPS is essential for genetic counselling. (2) Methods: this retrospective analysis focused on 2927 consecutive patients with WTs registered between 1989 and 2017 in the SIOP/GPOH studies. (3) Results: Genitourinary malformations (GU, = 66, 2.3%), Beckwith-Wiedemann spectrum (BWS, = 32, 1.1%), isolated hemihypertrophy (IHH, = 29, 1.0%), Denys-Drash syndrome (DDS, = 24, 0.8%) and WAGR syndrome ( = 20, 0.7%) were reported most frequently. Compared to others, these patients were younger at WT diagnosis (median age 24.5 months vs. 39.0 months), had smaller tumors (349.4 mL vs. 487.5 mL), less often metastasis (8.2% vs. 18%), but more often nephroblastomatosis (12.9% vs. 1.9%). WT with IHH was associated with blastemal WT and DDS with stromal subtype. Bilateral WTs were common in WAGR (30%), DDS (29%) and BWS (31%). Chemotherapy induced reduction in tumor volume was poor in DDS (0.4% increase) and favorable in BWS (86.9% reduction). The event-free survival (EFS) of patients with BWS was significantly ( = 0.002) worse than in others. (4) Conclusions: CPS should be considered in WTs with specific clinical features resulting in referral to a geneticist. Their outcome was not always favorable.
PubMed: 34638500
DOI: 10.3390/cancers13195016